Fig. 4

Up-regulation of hMT3 induces the remodeling of sorafenib-sensitive HCC tumors into sorafenib-resistant phenotype. A Representative 3D CLSM micrographs of untreated microtumors formed by Huh7^mock, Huh7^hMT3, and BCLC-3^WT cells. B Fluorescence micrographs of microtumors on the area of implantation of HCC cells on CAM either non-treated or treated with sorafenib (24 h). HCC cells are green (labeled with CellTracker Green), angiogenic blood vessels (BV) are red (labeled with rhodamine-LCA). Scale bars, 1 mm. In a zoomed micrographs, individual HCC cells that detached from the main microtumor could be detected intravasating into the endothelial cells in the adjacent tissues. Scale bars, 200 μm. C Quantification of the relative area (%) of the HCC microtumors and (D) invasive vasculotropic (intravasation, %) HCC cells that escaped from the primary tumor. Data are expressed as mean ± SEM. *p ≤ 0.05, **p ≤ 0.001. E Fluorescence micrographs of lungs excised from CAM assay. Metastasizing HCC cells are green, BV are red and nuclei blue (Hoechst 33258). Scale bars, 100 μm